Method of solubilizing and encapsulating itraconazole

ABSTRACT

Methods for preparing microspheres containing imidazole derivatives are provided. Also provided is the use of imidazole derivatives containing microspheres for treating fungal infections. Oral dosage forms for oral administration are also provided.

FIELD OF THE INVENTION

[0001] The present invention relates to the preparation of solutionscontaining imidazole derivatives and to the use of those solutions inthe preparation of microspheres. The imidazole derivative containingmicrospheres are effective in treating fungal infections, particularlyin mammals. The microspheres facilitate the oral administration ofrelatively large amounts of the imidazole derivative, with increasedbioavailability.

BACKGROUND OF THE INVENTION

[0002] Many present systems for delivering active agents to targets areseverely limited by biological, chemical, and physical barriers, whichare imposed by the environment through which delivery occurs, theenvironment of the target itself, or the target itself. Delivery is alsolimited, in many instances, by the chemical nature of the active agent.For example, oral delivery is generally ineffective with active agentsthat are poorly water-soluble.

[0003] The imidazole derivative family of compounds is particularlyeffective against a broad range of fungal infections such as thosecaused by Trichophyton rubrum, Tricophyton mentagrophytes,Epidermophyton flocesum, and Candida albicans, but these compounds areeither partially water soluble or insoluble in water. For example, thesolubility of itraconazole in water is less than 0.00001 g/ml.

[0004] Partially because imidazole derivatives are typically insolublein water, they are difficult to administer orally. Consequently althoughimidazole derivatives are frequently prescribed for the treatment offungal infections, they have been available only in topical preparationsor in oral formulations with limited bioavailability.

[0005] In recent years, fungal infections, such as those caused byCandida albicans in particular have become more prevalent andintractable due to their appearance in immunocompromised patients, suchas those infected with Human Immunodeficiency Virus (HIV) or thosesuffering from Acquired Immunodeficiency Syndrome (AIDS).

[0006] For example, U.S. Pat. No. 3,717,655 discloses imidazolederivatives which have antifungal and antibacterial activity. Thesecompounds are almost insoluble in aqueous solutions such as water andare very poorly soluble in polar solvents such as ethanol.

[0007] Das et al., U.S. Pat. No. 4,912,124, disclose a solvent systemfor imidazole derivatives that include mixtures of a polar solvent, apolyhydric alcohol that acts as a solubilizing agent, a nonionic oramphoteric surfactant, and a cosmetic humectant. Solutions containing atleast 1 percent by weight of the imidazole derivatives can be formulatedusing this solvent system. However, these formulations are suitable forexternal topical use only.

[0008] Accordingly, there is a need for orally deliverable forms ofimidazole derivative antifungal agents.

SUMMARY OF THE INVENTION

[0009] The present invention provides solutions comprising:

[0010] (a) at least about 2.5 parts by weight, based upon 100 parts byweight of solution, of a solute having the formula

[0011] wherein R, R¹, and R² are independently hydrogen or lower alkyl;

[0012] R³ is hydrogen, methyl or ethyl;

[0013] R⁴ is hydrogen or methyl

[0014] Ar is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orhalothienyl;

[0015] Ar¹ is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orcyanophenyl; and

[0016] n is 1 or 2; and

[0017] (b) a solubilizing effective amount of a solvent comprising atleast one volatile organic acid solvent.

[0018] Imidazole derivative microspheres are also provided. Thesemicrospheres comprise:

[0019] (a) an imidazole derivative active agent having the formula

[0020] wherein R, R¹, and R² are independently hydrogen or lower alkyl;

[0021] R³ is hydrogen, methyl or ethyl;

[0022] R⁴ is hydrogen or methyl

[0023] Ar is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orhalothienyl;

[0024] Ar¹ is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orcyanophenyl; and

[0025] n is 1 or 2; and

[0026] (b) a microsphere forming carrier selected from the groupconsisting of

[0027] (i) a proteinoid;

[0028] (ii) an acylated amino acid, poly amino acid, or a salt thereof;

[0029] (iii) an sulfonated amino acid, poly amino acid, or a saltthereof;

[0030] (iv) a protein or a salt thereof;

[0031] (v) an enteric coating material; or

[0032] (vi) any combination thereof.

[0033] Also contemplated by the present invention is a method forpreparing these microspheres. The method comprises:

[0034] (A) nebulizing a solution comprising

[0035] (a) an imidazole active agent having the formula

[0036] wherein R, R¹, and R² are independently hydrogen or lower alkyl;

[0037] R³ is hydrogen, methyl or ethyl;

[0038] R⁴ is hydrogen or methyl

[0039] Ar is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orhalothienyl;

[0040] Ar¹ is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orcyanophenyl; and

[0041] n is 1 or 2;

[0042] (b) an active agent and carrier solubilizing effective amount ofa solvent comprising an aqueous solution of at least one volatileorganic solvent; and wherein the volume:volume ratio of acid to water insaid carrier solution is at least about 3:7, and

[0043] (c) microsphere forming a carrier selected from the groupconsisting of

[0044] (i) a proteinoid;

[0045] (ii) an acylated amino acid or poly amino acid or a salt thereof;

[0046] (iii) an sulfonated amino acid or poly amino acid or a saltthereof;

[0047] (iv) a protein or a salt thereof;

[0048] (v) an enteric coating material; or

[0049] (vi) any combination thereof; and

[0050] (B) decreasing said ratio to less than about 3:7, to yield saidmicrospheres. Alternatively, the active agent and the carrier can besolubilized in separate solutions. The separate solutions can benebulized together and the acid to water ratio then decreased as above.

[0051] Methods for the oral administration of imidazole derivatives arealso contemplated wherein the microsphere compositions above are orallyadministered to an animal in need of this treatment.

BRIEF DESCRIPTION OF THE DRAWINGS

[0052] FIGS. 1A-1H are scanning electron microspheres of itraconazolecontaining microspheres prepared according to the present invention.

DETAILED DESCRIPTION OF THE INVENTION

[0053] It has now been discovered that water insoluble or partiallysoluble imidazole derivatives can be solubilized in volatile organicacids. The resultant solutions can be used to prepare imidazolecontaining microspheres which are suitable for oral administration toanimals.

[0054] Imidazole Derivatives

[0055] The active agents of the present invention are imidazolederivatives having the formula:

[0056] wherein R, R¹ and R² are independently hydrogen or lower alkyl;

[0057] R³ is hydrogen, methyl or ethyl;

[0058] R⁴ is hydrogen or methyl

[0059] Ar is phenyl, monohalophenyl, dihalophenyl, trihalophenyl, mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orhalothienyl;

[0060] Ar¹ is phenyl, monohalophenyl, dihalophenyl, trihalophenyl,mono(lower alkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, orcyanophenyl; and

[0061] n is 1 or 2.

[0062] A preferred imidazole derivative is itraconazole. Itraconazole isa synthetic triazole imidazole derivative 1:1:1:1 racemic mixture offour diastereomers (two enantiomeric pairs), each possessing threechiral centers (Physicians Desk Reference 48th Ed., pg. 1097, 1994).

[0063] Imidazole Derivative Solutions

[0064] The solutions prepared in accordance with the present inventionallow for the solubilization of imidazole derivatives at concentrationssuitable for processing into orally administrable forms havingacceptable bioavailability.

[0065] In accordance with the present invention, imidazole derivativesare solubilized in volatile organic acid solvent(s). Preferred acidsolvents for the imidazole derivatives are acetic acid and formic acid.Preferably, the solvent itself is an aqueous solution of the acid. Mostpreferably the volume:volume ratio of the acid to the total volume ofthe solvent is 3:7 or greater. It has been found that by using thissolvent system up to a 50% solution of imidazole derivative can beprepared.

[0066] Dissolution is achieved by simple mixing, with heating ifnecessary. The more concentrated the acid in the solvent, the greaterthe amount of active agent that can be incorporated into the solution.If lower concentrations of acid are required for the end use of thesolution, the active agent can first be dissolved in a more concentratedacid solution, and the resultant solution then slowly diluted further,preferably with water.

[0067] Preferably, the solution comprises from about 3 to about 40percent by weight of solute and from about 60 to about 97 parts byweight of solvent based upon 100 parts by weight of solution.

[0068] The solvent itself, preferably comprises from about 30 to about80 parts by volume of acid and from about 70 to about 20 parts by volumeof water based upon 100 parts by volume of solvent. Most preferably, thesolvent comprises from about 40 to about 50 parts by volume of acid andfrom about 60 to about 50 parts by volume of water based upon 100 partsby volume of solvent.

[0069] Microspheres

[0070] Microspheres are useful in the delivery of active agents becausethey protect an active agent cargo until it is delivered to a target.Microspheres are particularly useful in the oral delivery ofbiologically active agents such as, for example, pharmaceutically activeagents. Copending Application entitled “Method of Solubilizing andEncapsulating Itraconazole”, filed on [. . . ] discloses methods andapparatus for forming microspheres.

[0071] Microspheres containing an active agent can be generally of thematrix form or the capsule form. In a hollow matrix spheroid form, thecenter of the sphere is hollow and the cargo or active agent isdistributed throughout a carrier matrix. In a solid matrix form, thecarrier matrix forms a continuum in which the cargo is distributed. Inthe microcapsule form, the encapsulated material or cargo can be eitherin solution or a solid, with the carrier forming a shell around thecargo.

[0072] The methods of the present invention are cost-effective forpreparing microspheres which contain imidazole derivatives, are simpleto perform, and are amenable to industrial scale-up for commercialproduction.

[0073] Carriers

[0074] Carriers suitable for use in the present invention aremicrosphere forming carriers. These carriers include, withoutlimitation, proteinoids; acylated amino acids, poly amino acids or saltsthereof; sulfonated amino acids, poly amino acids or salts thereof;proteins or salts thereof, enteric coating materials; or any combinationthereof.

[0075] Amino acids are the basic materials used to prepare many of thecarriers useful in the present invention. Amino acids include anycarboxylic acid having at least one free amino group and includenaturally occurring and synthetic amino acids. The preferred amino acidsfor use in the present invention are ∝-amino acids and, most preferably,are naturally occurring ∝-amino acids. Many amino acids and amino acidesters are readily available from a number of commercial sources such asAldrich Chemical Co. (Milwaukee, Wis., USA); Sigma Chemical Co. (St.Louis, Mo., USA); and Fluka Chemical Corp. (Ronkonkoma, N.Y., USA).

[0076] Representative, but not limiting, amino acids suitable for use inthe present invention are generally of the formula

[0077] wherein:

[0078] R⁵ is hydrogen, C₁-C₄ alkyl, or C₂-C₄ alkenyl;

[0079] R⁶ is C₁-C₂₄ alkyl, C₂-C₂₄ alkenyl, C₃-C₁₀ cycloalkyl, C₃-C₁₀cycloalkenyl, phenyl, naphthyl, (C₁-C₁₀ alkyl) phenyl, (C₂-C₁₀ alkenyl)phenyl, (C₁-C₁₀ alkyl) naphthyl, (C₂-C₁₀ alkenyl) naphthyl, phenyl(C₁-C₁₀ alkyl), phenyl (C₂-C₁₀ alkenyl), naphthyl (C₁-C₁₀ alkyl), ornaphthyl (C₂-C₁₀ alkenyl);

[0080] R⁶ being optionally substituted with C₁-C₄ alkyl, C₂-C₄ alkenyl,C₁-C₄ alkoxy, —OH, —SH, —CO₂R⁷, C₃-C₁₀ cycloalkyl, C₃-C₁₀ cycloalkenyl,heterocycle having 3-10 ring atoms wherein the hetero atom is one ormore of N, O, S, or any combination thereof, aryl, (C₁-C₁₀ alk)aryl,ar(C₁-C₁₀ alkyl) or any combination thereof;

[0081] R⁶ being optionally interrupted by oxygen, nitrogen, sulfur, orany combination thereof; and

[0082] R⁷ is hydrogen, C₁-C₄ alkyl, or C₂-C₄ alkenyl.

[0083] The preferred naturally occurring amino acids for use in thepresent invention as amino acids or components of a peptide are alanine,arginine, asparagine, aspartic acid, citrulline cysteine, cystine,glutamic acid, glutamine, glycine, histidine, isoleucine, leucine,lysine, methionine, ornithine, phenylalanine, proline, serine,threonine, tryptophan, tyrosine, valine, hydroxyproline,β-carboxyglutamic acid, γ-carboxyglutamic acid, phenylglycine, orO-phosphoserine. The most preferred amino acids are arginine, leucine,lysine, phenylalanine, tyrosine, tryptophan, valine, and phenylglycine.

[0084] The preferred non-naturally occurring amino acids for use in thepresent invention are β-alanine, α-amino butyric acid, γ-amino butyricacid, γ-(aminophenyl) butyric acid, α-amino isobutyric acid, ε-aminocaproic acid, 7-amino heptanoic acid, β-aspartic acid, aminobenzoicacid, aminophenyl acetic acid, aminophenyl butyric acid, γ-glutamicacid, cysteine (ACM), ε-lysine, methionine sulfone, norleucine,norvaline, ornithine, d-ornithine, p-nitro-phenylalanine, hydroxyproline, 1,2,3,4,-tetrahydroisoquinoline-3-carboxylic acid, andthioproline.

[0085] Poly amino acids are either peptides or two or more amino acidslinked by a bond formed by other groups which can be linked, e.g., anester or an anhydride linkage. Special mention is made of non-naturallyoccurring poly amino acids and particularly non-naturally occurringhetero-poly amino acids, i.e. of mixed amino acids.

[0086] Peptides are two or more amino acids joined by a peptide bond.Peptides can vary in length from di-peptides with two amino acids topolypeptides with several hundred amino acids. See, Walker, ChambersBiological Dictionary, Cambridge, England: Chambers Cambridge, 1989,page 215. Special mention is made of non-naturally occurring peptidesand particularly non-naturally occurring peptides of mixed amino acids.Special mention is also made of di-peptides tri-peptides,tetra-peptides, and penta-peptides, and particularly, the preferredpeptides are di-peptides and tri-peptides. Peptides can be homo- orhetero- peptides and can include natural amino acids, synthetic aminoacids, or any combination thereof.

[0087] Proteinoids

[0088] Proteinoids are artificial polymers of amino acids. Proteinoidspreferably are prepared from mixtures of amino acids. Preferredproteinoids are condensation polymers, and most preferably, are thermalcondensation polymers. These polymers may be directed or randompolymers. Proteinoids can be linear, branched, or cyclical, and certainproteinoids can be units of other linear, branched, or cyclicalproteinoids.

[0089] Special mention is made of diketopiperazines. Diketopiperazinesare six member ring compounds. The ring includes two nitrogen atoms andis substituted at two carbons with two oxygen atoms. Preferably, thecarbonyl groups are at the 2 and 5 ring positions. These rings can beoptionally, and most often are, further substituted.

[0090] Diketopiperazine ring systems may be generated during thermalpolymerization or condensation of amino acids or amino acid derivatives.(Gyore, J; Ecet M. Proceedings Fourth ICTA (Thermal Analysis), 1974, 2,387-394 (1974)). These six membered ring systems were presumablygenerated by intra-molecular cyclization of the dimer prior to furtherchain growth or directly from a linear peptide (Reddy, A. V., Int. J.Peptide Protein Res., 40, 472-476 (1992); Mazurov, A. A. et al., Int. J.Peptide Protein Res., 42, 14-19 (1993)).

[0091] Diketopiperazines can also be formed by cyclodimerization ofamino acid ester derivatives as described by Katchalski et al., J. Amer.Chem. Soc., 68, 879-880 (1946), by cyclization of dipeptide esterderivatives, or by thermal dehydration of amino acid derivatives andhigh boiling solvents as described by Kopple et al., J. Org. Chem., 33(2), 862-864 (1968).

[0092] Diketopiperazines typically are formed from α-amino acids.Preferably, the α-amino acids of which the diketopiperazines are derivedare glutamic acid, aspartic acid, tyrosine, phenylalanine, and opticalisomers of any of the foregoing.

[0093] Modified Amino Acids and Poly Amino Acids

[0094] Modified amino acids, poly amino acids, or peptides are eitheracylated or sulfonated and include amino acid amides and sulfonamides.

[0095] Acylated Amino Acids and Poly Amino Acids

[0096] Although any acylated amino acids or poly amino acids are usefulin the present invention, special mention is made of acylated aminoacids having the formula

Ar²—Y—(R⁸)_(n)—OH  III

[0097] wherein Ar² is a substituted or unsubstituted phenyl or naphthyl;

[0098] Y is

[0099] R⁸ has the formula

[0100]  wherein:

[0101] R⁹ is C₁ to C₂₄ alkyl, C₁ to C₂₄ alkenyl, phenyl, naphthyl, (C₁to C₁₀ alkyl) phenyl, (C₁ to C₁₀ alkenyl) phenyl, (C₁ to C₁₀ alkyl)naphthyl, (C₁ to C₁₀ alkenyl) naphthyl, phenyl (C₁ to C₁₀ alkyl), phenyl(C₁ to C₁₀ alkenyl), naphthyl (C₁ to C₁₀ alkyl) and naphthyl (C₁ to C₁₀alkenyl);

[0102] R⁹ is optionally substituted with C₁ to C₄ alkyl, C₁ to C₄alkenyl, C₁ to C₄ alkoxy, —OH, —SH and —CO₂R¹¹, cycloalkyl,cycloalkenyl, heterocyclic alkyl, alkaryl, heteroaryl, heteroalkaryl, orany combination thereof;

[0103] R¹¹ is hydrogen, C₁ to C₄ alkyl or C₁ to C₄ alkenyl;

[0104] R⁹ is optionally interrupted by oxygen, nitrogen, sulfur or anycombination thereof; and

[0105] R¹⁰ is hydrogen, C₁ to C₄ alkyl or C₁ to C₄ alkenyl.

[0106] Special mention is also made of those having the formula

[0107] wherein:

[0108] R¹² is (i) C₃-C₁₀ cycloalkyl, optionally substituted with C₁-C₇alkyl, C₂-C₇ alkenyl, C₁-C₇ alkoxy, hydroxy, phenyl, phenoxy or —CO₂R¹⁵,wherein R¹⁵ is hydrogen, C₁-C₄ alkyl, or C₂-C₄ alkenyl; or

[0109] (ii) C₁-C₆ alkyl substituted with C₃-C₁₀ cycloalkyl;

[0110] R¹³ is hydrogen, C₁-C₄ alkyl, or C₂-C₄ alkenyl;

[0111] R¹⁴ is C₁-C₂₄ alkyl, C₂-C₂₄ alkenyl, C₃-C₁₀ cycloalkyl, C₃-C₁₀cycloalkenyl, phenyl, naphthyl, (C₁-C₁₀ alkyl) phenyl, (C₂-C₁₀ alkenyl)phenyl, (C₁-C₁₀ alkyl) naphthyl, (C₂-C₁₀ alkenyl) naphthyl, phenyl(C₁-C₁₀ alkyl), phenyl (C₂-C₁₀ alkenyl), naphthyl (C₁-C₁₀ alkyl) ornaphthyl (C₂-C₁₀ alkenyl);

[0112] R¹⁴ being optionally substituted with C₁-C₄ alkyl, C₂-C₄ alkenyl,C₁-C₄ alkoxy, —OH, —SH, —CO₂R¹⁶, C₃-C₁₀ cycloalkyl, C₃-C₁₀ cycloalkenyl,heterocycle having 3-10 ring atoms wherein the hetero atom is one ormore of N, O, S or any combination thereof, aryl, (C₁-C₁₀ alk)aryl,ar(C₁-C₁₀ alkyl), or any combination thereof;

[0113] R¹⁴ being optionally interrupted by oxygen, nitrogen, sulfur, orany combination thereof; and

[0114] R¹⁶ is hydrogen, C₁-C₄ alkyl, or C₂-C₄ alkenyl.

[0115] Acylated amino acids may be prepared by reacting single aminoacids, mixtures of two or more amino acids, or amino acid esters with anamine modifying agent which reacts with free amino moieties present inthe amino acids to form amides.

[0116] Suitable, but non-limiting, examples of acylating agents usefulin preparing acylated amino acids include

[0117] acid chloride acylating agents having the formula

[0118]  wherein:

[0119] R¹⁷ an appropriate group for the modified amino acid beingprepared, such as, but not limited to, alkyl, alkenyl, cycloalkyl, oraromatic, and particularly methyl, ethyl, cyclohexyl, cyclophenyl,phenyl, or benzyl, and X is a leaving group. Typical leaving groupsinclude, but are not limited to, halogens such as chlorine, bromine andiodine.

[0120] Examples of the acylating agents include, but are not limited to,acyl halides including, but not limited to, acetyl chloride, propylchloride, cyclohexanoyl chloride, cyclopentanoyl chloride, andcycloheptanoyl chloride, benzoyl chloride, hippuryl chloride and thelike; and anhydrides, such as acetic anhydride, propyl anhydride,cyclohexanoic anhydride, benzoic anhydride, hippuric anhydride and thelike. Preferred acylating agents include benzoyl chloride, hippurylchloride, acetyl chloride, cyclohexanoyl chloride, cyclopentanoylchloride, and cycloheptanoyl chloride.

[0121] The amine groups can also be modified by the reaction of acarboxylic acid with coupling agents such as the carbodiimidederivatives of amino acids, particularly hydrophilic amino acids such asphenylalanine, tryptophan, and tyrosine. Further examples includedicyclohexylcarbodiimide and the like.

[0122] If the amino acid is multifunctional, i.e. has more than one —OH,—NH₂ or —SH group, then it may optionally be acylated at one or morefunctional groups to form, for example, an ester, amide, or thioesterlinkage.

[0123] In acylated poly amino acids, one or more of the amino acids maybe modified (acylated). Modified poly amino acids may include one ormore acylated amino acid(s). Although linear modified poly amino acidswill generally include only one acylated amino acid, other poly aminoacid configurations can include more than one acylated amino acid. Polyamino acids can be polymerized with the acylated amino acid(s) or can beacylated after polymerization.

[0124] Sulfonated Amino Acids and Poly Amino Acids

[0125] Sulfonated amino acids and poly amino acids are modified bysulfonating at least one free amine group with a sulfonating agent whichreacts with at least one of the free amine groups present.

[0126] Special mention is made of compounds of the formula

Ar³—Y—(R¹⁸)_(n)—OH  V

[0127] wherein Ar³ is a substituted or unsubstituted phenyl or naphthyl;

[0128] Y is —SO₂—, R¹⁸ has the formula

[0129]  wherein:

[0130] R¹⁹ is C₁ to C₂₄ alkyl, C₁ to C₂₄ alkenyl, phenyl, naphthyl, (C₁to C₁₀ alkyl) phenyl, (C₁ to C₁₀ alkenyl) phenyl, (C₁ to C₁₀ alkyl)naphthyl, (C₁ to C₁₀ alkenyl) naphthyl, phenyl (C₁ to C₁₀ alkyl), phenyl(C₁ to C₄ alkenyl), naphthyl (C₁ to C₁₀ alkyl) and naphthyl (C₁ to C₁₀alkenyl);

[0131] R¹⁹ is optionally substituted with C₁ to C₄ alkyl, C₁ to C₄alkenyl, C₁ to C₄ alkoxy, —OH, —SH and —CO₂R²¹ or any combinationthereof;

[0132] R²¹ is hydrogen, C₁ to C₄ alkyl or C₁ to C₄ alkenyl;

[0133] R¹⁹ is optionally interrupted by oxygen, nitrogen, sulfur or anycombination thereof; and

[0134] R²⁰ is hydrogen, C₁ to C₄ alkyl or C₁ to C₄ alkenyl.

[0135] Suitable, but non-limiting, examples of sulfonating agents usefulin preparing sulfonated amino acids include sulfonating agents havingthe formula R²²—So₂—X wherein R²² is an appropriate group for themodified amino acid being prepared such as, but not limited to, alkyl,alkenyl, cycloalkyl, or aromatics and X is a leaving group as describedabove. One example of a sulfonating agent is benzene sulfonyl chloride.

[0136] Modified poly amino acids and peptides may include one or moresulfonated amino acid(s). Although linear modified poly amino acids andpeptides used generally include only one sulfonated amino acid, otherpoly amino acid and peptide configurations can include more than onesulfonated amino acid. Poly amino acids and peptides can be polymerizedwith the sulfonated amino acid(s) or can be sulfonated afterpolymerization.

[0137] Proteins

[0138] Proteins are naturally occurring (i.e. not artificial) polymersof amino acids.

[0139] Enteric Coating Materials

[0140] Enteric coating materials known to those skilled in the art suchas, for example, cellulose acetate trimellitate (CAT) and celluloseacetate phthalate (CAP), are suitable for use in the preservation aswell.

[0141] Formation

[0142] These carriers, and particularly proteinoids, acylated aminoacids or poly amino acids, sulfonated amino acids or poly amino acids,and proteins are often insoluble or relatively insoluble in neutral ormildly acidic solutions but are also soluble, as are the imidazolederivatives useful in the present invention, in aqueous acid solutionswherein the volume to volume ratio of acid to water is greater thanabout 3:7. Suitable aqueous acid solvents are as above, i.e. volatileorganic acids, such as for example, aqueous acetic acid, aqueous formicacid, and the like. These acids will volatilize upon nebulization or canbe diluted in the aqueous solution, thereby decreasing the concentrationof the acid and reversing the solubility of the carrier even in theabsence of a precipitator. For example, see currently filed U.S. patentapplication Ser. No. ______ , (attorney's docket no. 1946/09202)entitled “SPRAY DRYING METHOD AND APPARATUS”.

[0143] Microsphere formation occurs when the concentration of the acidin the carrier/active agent solution is decreased. As this solution isnebulized, the acid evaporates, decreasing the concentration of the acidin solution to less than 30% by volume. The carrier, then, will selfassemble to form microspheres containing any optional active agent. Thecargo must be stable in the concentrated acid for the time andconditions necessary to carry out the operation. Alternately, thecarrier solution can be diluted, such as with water, whereby the acidconcentration is decreased and the carrier precipitates to formmicrospheres. Preferably, the microspheres are prepared by spray drying.

[0144] The microspheres can be pH adapted by using base or acid solublecoatings including, but not limited to, proteinoid coatings, entericcoatings, acylated amino acid coatings, and the like.

[0145] Any of the solutions above may optionally contain additives suchas stabilizing additives. The presence of such additives promotes thestability and dispersability of the active agent in solution. Thestabilizing additives may be employed at a concentration ranging betweenabout 0.1 and 5% (w/v), preferably about 0.5% (w/v). Suitable, butnon-limiting examples of stabilizing additives include buffer salts, gumacacia, gelatin, methyl cellulose, polyethylene glycol, and polylysine.

[0146] The amount of active agent that may be incorporated in themicrosphere is dependent upon a number of factors which include theconcentration of active agent in the solution as well as the affinity ofthe active agent for the carrier. The concentration of the active agentin the final formulation also will vary depending on the requiredamounts for any particular end use. When necessary, the exactconcentration can be determined by, for example, reverse phase HPLCanalysis.

[0147] The microspheres and, therefore, the solutions described abovemay also include one or more enzyme inhibitors. Such enzyme inhibitorsinclude, but are not limited to, compounds such as actinonin orepiactinonin and derivatives thereof.

[0148] The microspheres are particularly useful for administeringitraconazole derivatives to any animals, including but not limited to,birds and mammals, such as primates and particularly humans; andinsects. These microsphere systems are particularly advantageous fordelivering these active agents as the active agent would otherwise bedestroyed or rendered less effective by conditions encountered beforethe microsphere reaches the active agent target zone (i.e., the area inwhich the active agent of the delivery composition are to be released)and within the body of the animal to which they are administered.Furthermore, these microspheres can deliver relatively high amounts ofthe imidazole derivative and retain a high bioavailability.

Description of the Preferred Embodiments

[0149] The following examples illustrate the present invention withoutlimitation.

EXAMPLE 1 Solubilization of Itraconazole

[0150] Acetic acid solutions were prepared in water to 10%, 20%, 50% and75% concentrations (expressed as volume glacial acetic acid/total volumeof solution×100). 100 mg itraconazole solute were then mixedindependently with 1 ml of each solution and visually monitored fordissolution. If necessary, additional 1 ml aliquots of each acetic acidsolution were added until the itraconazole solute was dissolved.

[0151] Results are illustrated in Table 1 below. The solubilizedmaterial did not precipitate readily. TABLE 1 SOLUBILITY ConcentrationAmount of Solvent of Acid 1 ml 1 ml 2 ml 2 ml 3 ml 3 ml 4 ml 4 ml inSolvent Cold Hot Cold Hot Cold Hot Cold Hot 10% Acetic Ins. Ins. Ins.Ins. Ins. Ins. — Ins. Acid v:v 20% Acetic Ins. Ins. Ins. Ins. Ins. Part— — Acid v:v Sol. 50% Acetic Ins. Ins. Ins. Sol. Ins. — — — Acid v:v 75%Acetic Sol. — — — — — — — Acid v:v

EXAMPLE2 Solubilization of Itraconazole

[0152] 100 mg of itraconazole solute were dissolved in 1 ml glacialacetic acid solvent and aqueous acetic acid solvent at variousconcentrations. Results are illustrated in Table 2 below. TABLE 2ITRACONAZOLE SOLUTE % Acetic Acid (v:v) Dissolved Itraconazole % 100 >33(dissolves freely on addition) 75 >10 (dissolves freely on addition) 405 (diss.conc.acid, then dilute) 20 2.5 (diss.conc.acid, then dilute)

EXAMPLE 3 Preparation of Itraconazole-Containing MicrospheresOne-Solution Method

[0153] 60 grams of itraconazole solute (Janssen Pharmaceutica) wereadded to 1.43 liters of glacial acetic acid solvent, and the mixture wasstirred to dissolve the solute. 1.43 liters of water were then addedusing a pump at a flow rate of 25 ml/min. Slight clouding of thesolution was observed, but cleared upon further stirring. 166 grams ofproteinoid (Glu-Asp-Tyr-Phe-Orn) were added and dissolved with furtherstirring. The final solution was filtered through folded tissue paper.

[0154] Using peristaltic pumps, the solution was fed through a VirtisSD04 spray drying apparatus under the conditions of Table 3 below. TABLE3 SPRAY DRYING CONDITIONS Solution flow rate 7-8 ml/min Inlettemperature 175° C. Outlet temperature 116° C. Blower speed fullCompressor pressure full

[0155] Stable proteinoid microspheres containing itraconazole wereformed. Analysis of typical microspheres using RP-HPLC demonstrated thatthey contained 14-21% itraconazole by weight.

[0156] Scanning electron microscopy in FIGS. 1A-1H illustrates that themicrospheres were smooth and spherical and had diameters ranging from0.1 μm to about 5 μm. When mechanically crushed only the larger spheresshattered, while the smaller spheres remained intact. Crushing revealeda solid internal structure. See, FIGS. 1G and 1H.

[0157] All patents, applications, publications, and test methodsmentioned herein are hereby incorporated by reference in their entirety.

[0158] Many variations of the present invention will suggest themselvesto those skilled in the art in light of the above-detailed descriptionin which obvious variations are within the full intended scope of theappended claims.

What is claimed is:
 1. A solution comprising: (a) at least about 2.5parts by weight, based upon 100 parts by weight of solution, of a solutehaving the formula

wherein R, R¹, and R ² are independently hydrogen or lower alkyl; R³ ishydrogen, methyl or ethyl; R⁴ is hydrogen or methyl Ar is phenyl,monohalophenyl, dihalophenyl, trihalophenyl, mono(lower alkyl)phenyl,di(lower alkyl)phenyl, lower alkoxyphenyl, or halothienyl; Ar¹ isphenyl, monohalophenyl, dihalophenyl, trihalophenyl, mono(loweralkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, or cyanophenyl;and n is 1 or 2; and (b) a solubilizing effective amount of a solventcomprising at least one volatile organic acid.
 2. A solution as definedin claim 1, wherein said solute comprises itraconazole.
 3. A solution asdefined in claim 1, wherein said solvent comprises an aqueous solutionof said acid.
 4. A solution as defined in claim 3, wherein said solventcomprises aqueous acetic acid.
 5. A solution is defined as in claim 3,wherein said solvent comprises aqueous formic acid.
 6. A solution isdefined in claim 1, wherein said solution comprises from about 3 toabout 40 percent by weight of solute and from about 60 to about 97 partsby weight of solvent based upon 100 parts by weight of solution.
 7. Asolution as defined in claim 3, wherein said solvent comprises fromabout 30 to about 80 parts by volume of acid and from about 70 to about20 parts by volume of water, based upon 100 parts by volume of solvent.8. A solution as defined in claim 7, wherein said solvent comprises fromabout 40 to about 50 parts by volume weight of acid and from about 60 toabout 50 parts by volume of water, based upon 100 parts by volume ofsolvent.
 9. A solution is defined in claim 3, wherein the volume:volumeratio of acid to water in said solvent is at least about 3:7.
 10. Amicrosphere comprising (a) an imidazole derivative active agent havingthe formula

wherein R, R¹, and R² are independently hydrogen or lower alkyl; R³ ishydrogen, methyl or ethyl; R⁴ is hydrogen or methyl Ar is phenyl,monohalophenyl, dihalophenyl, trihalophenyl, mono(lower alkyl)phenyl,di(lower alkyl)phenyl, lower alkoxyphenyl, or halothienyl; Ar¹ isphenyl, monohalophenyl, dihalophenyl, trihalophenyl, mono(loweralkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, or cyanophenyl;and n is 1 or 2; and (b) a microsphere forming carrier selected from thegroup consisting of (i) a proteinoid; (ii) an acylated amino acid orpoly amino acid or a salt thereof; (iii) a sulfonated amino acid or polyamino acid or a salt thereof; (iv) a protein or a salt thereof; (v) anenteric coating material; or (vi) any combination thereof.
 11. Amicrosphere as defined in claim 10, wherein said imidazole derivativeactive agent comprises itraconazole.
 12. A method for preparingimidazole derivative active agent containing microspheres, said methodcomprising (A) nebulizing a solution comprising (a) an imidazole activeagent having the formula

wherein R, R¹, and R² are independently hydrogen or lower alkyl; R³ ishydrogen, methyl or ethyl; R⁴ is hydrogen or methyl Ar is phenyl,monohalophenyl, dihalophenyl, trihalophenyl, mono(lower alkyl)phenyl,di(lower alkyl)phenyl, lower alkoxyphenyl, or halothienyl; Ar¹ isphenyl, monohalophenyl, dihalophenyl, trihalophenyl, mono(loweralkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, or cyanophenyl;and n is 1 or 2; (b) a microsphere forming carrier selected from thegroup consisting of (i) a proteinoid; (ii) an acylated amino acid orpoly amino acid or a salt thereof; (iii) a sulfonated amino acid or polyamino acid or a salt thereof; (iv) a protein or a salt thereof; (v) anenteric coating material; or (vi) any combination thereof; and (c) anactive agent and carrier solubilizing effective amount of a solventcomprising an aqueous solution of at least one volatile organic acid;wherein the volume:volume ratio of acid to water in said solvent is atleast about 3:7; and (B) decreasing said ratio to less than about 3:7,to yield said microspheres.
 13. A method as defined in claim 12, whereinsaid imidazole derivative active agent comprises itraconazole.
 14. Amethod as defined in claim 12, wherein said aqueous solution comprisesaqueous acetic acid.
 15. A method as defined in claim 12, wherein saidnebulizing is performed by spray drying.
 16. A method wherein said ratiois decreased by volatilizing at least a portion of said acid.
 17. Amethod wherein said ratio is decreased by diluting said acid.
 18. Amethod for preparing imidazole derivative active agent containingmicrospheres, said method comprising (A) nebulizing (a) a solutioncomprising (i) an imidazole active agent having the formula

wherein R, R¹, and R² are independently hydrogen or lower alkyl; R³ ishydrogen, methyl or ethyl; R⁴ is hydrogen or methyl Ar is phenyl,monohalophenyl, dihalophenyl, trihalophenyl, mono(lower alkyl)phenyl,di(lower alkyl)phenyl, lower alkoxyphenyl, or halothienyl; Ar¹ isphenyl, monohalophenyl, dihalophenyl, trihalophenyl, mono(loweralkyl)phenyl, di(lower alkyl)phenyl, lower alkoxyphenyl, or cyanophenyl;and n is 1 or 2; and (ii) an active agent solubilizing effective amountof a solvent comprising an aqueous solution of at least one volatileorganic acid; wherein the volume:volume ratio of acid to water in saidsolvent is at least about 3:7; and (b) a solution of a microsphereforming carrier selected from the group consisting of (i) a proteinoid;(ii) an acylated amino acid or poly amino acid or a salt thereof; (iii)a sulfonated amino acid or poly amino acid or a salt thereof; (iv) aprotein or a salt thereof; (v) an enteric coating material; or (vi) anycombination thereof; and (B) decreasing said ratio to less than about3:7, to yield said microspheres.
 19. A method for administeringitraconazole to an animal in need of such treatment, such methodcomprising (A) preparing a composition as defined in claim 10, and (B)orally administering said composition to said animal.
 20. A method fortreating a fungal infection in an animal in need of such treatment, saidmethod comprising administering to said animal a therapeuticallyeffective amount of a composition as defined in claim 10.